The proposed research will be a systematic biochemical investigation of the mechanism of E. coli RNA Polymerase. The rate determining steps in RNA chain initiation are being studied employing kinetic, product analysis, and spectroscopic (fluorescence and absorption changes) techniques. The role of sigma subunit in promoter function will be investigated. The role of bacteriophage lambda N-protein in RNA chain antitermination will be studied. The mechanism of action of certain antibiotic inhibitors of RNA synthesis will be studied. The effect of positive control factors such as lambda repressor and CRP plus cAMP in promoting initiation from the lambda PRM and E. coli lac promoters will also be investigated. The mechanistic work is seen as a foundation on which the regulatory studies can most profitably be pursued. The characterization of promoter mutations at a detailed mechanistic level will allow a more thorough understanding of the biochemical bases of gene expression.